1. Field of the Invention
The present embodiments relates to systems and methods for synthesizing molecules such as polymers. More specifically, the present embodiments relate to devices and methods for continuously synthesizing biological polymers such as polypeptides and oligonucleotides.
2. Description of the Related Art
Oligonucleotides are among the most important and prevalent reagents used in biotechnology laboratories engaged in research, diagnostics and therapeutics. The high demand for oligonucleotides derives from their exquisite specificity for complementary nucleotide sequences in DNA or RNA obtained from biological samples. This specificity allows oligonucleotides to be used as probes that specifically bind to unique sequences present in less than part-per-billion abundance in a complex biological milieu. This specificity can be used in order to provide a diagnosis for an individual at risk for a particular disease based on the presence or absence of a gene sequence known to be associated with the disease. Furthermore, this specificity also forms the basis for use of oligonucleotides as reagents for synthesizing molecules of DNA or RNA having a particular nucleotide sequence of interest. For example, a gene sequence associated with a particular disease can be cloned by binding one or more oligonucleotides to the gene sequence and then performing an amplification reaction on the bound complex to make multiple copies of the sequence. The cloned gene sequence can subsequently be utilized for research into the disease or can even be used for therapeutic treatment of individuals afflicted with the disease.
Oligonucleotide synthesis is a cyclical process that assembles a chain of nucleotides. Nucleotides are added one by one through a cycle of chemical reactions, in which a particular molecule (e.g., a nucleotide) is added to a growing DNA molecule (e.g., a growing DNA chain), sometimes via catalysis, until the desired chain is complete. Generally, each cycle of chemical reactions includes the steps of detritylation, coupling, capping and oxidation. During the detritylation or “deprotection” step, a dimethoxytrityl (DMT) group is removed from the last nucleotide of the growing DNA chain to allow the addition of the next nucleotide. The amount of DMT released from each cycle is monitored to determine coupling efficiency. The release of DMT is apparent because a bright orange color is emitted as DMT is released.
Similar synthetic methods can be used to produce oligonucleotides at a variety of throughputs to satisfy demands spanning those of small laboratories to large manufacturing facilities. The methods themselves are relatively robust being capable of handling oligonucleotides of varying length from just a few nucleotides per molecule to over 100 nucleotides per molecule. Furthermore, the methods are capable of producing oligonucleotides having a myriad number of different sequences, the complexity of which is illustrated by the fact that the number of different decamers (molecules having 10 nucleotides) that can be made using just the 4 common DNA nucleotides (A, T, C and G) is 410=1,048,576.
Typically, large manufacturing facilities achieve high throughput by employing a large number of synthesizers that are similar to those used in smaller laboratories. These synthesizers are typically configured to perform individual steps of the monomer addition cycle in succession, and can do so for several different oligonucleotides in parallel. Thus, the sequence of reactions for a plurality of oligonucleotides is performed in order such that detritylation is carried out for each oligonucleotide, then coupling is carried out for each oligonucleotide, followed by capping of each oligonucleotide followed by oxidation of each oligonucleotide. The cycle is then repeated until full length oligonucleotides are obtained.
Although a large number of such synthesizers can be employed to achieve relatively high throughput, this scale-up approach can result in unwanted inefficiencies. An example of such inefficiency is the time that each synthesizer sits idle while waiting for individual incubation steps to be completed prior to delivery of the next reagent to any of the reactions. Furthermore, instruments can experience substantial down time between synthesis reactions when new reaction vessels or reagents are reloaded.
What is needed are synthetic methods and devices that allow parallel synthesis of oligonucleotides and other molecules such that a plurality of reactions can be carried out and monitored without interruption. The present embodiments satisfy this need and provides other advantages as well.